Detection Soil is one of the most

Detection of drugs in soil using HPLCAbstractSoil is one of the most valuable sources of trace evidence within forensic investigations, due to the transferability of the soil itself, and the variation in the heterogeneity and complexity of the soil. Soil is a common factor of trace evidence due to its persistence within the environment, since it can be found almost anywhere around the country. This kind of earth material can provide valuable information within forensic science to aid with investigations such as the geographical location that the soil sample was recovered from, whilst also allowing crime scene samples to be compared to control samples, and any samples that are not important to the investigation can be excluded. Previous studies have shown that there are major differences between forensic science in comparison with earth sciences, in particularly, the analysis and detection of drugs within soil samples. This study expresses an enhanced analytical technique for the detection of drugs in soil, based on previous studies of the analysis of soil via the use of high performance liquid chromatography, or HPLC for short.

This study yields information in the form of HPLC chromatograms, with clear indications within peak retention times that drugs can be detected in soil. The results attained from the HPLC of four blank samples of soil, each recovered from four separate locations, were compared with four spiked soil samples, which were also recovered from these four locations. The results therefore show that HPLC prospective to present a valuable, precise analytical method for the detection and analysis of drugs within soil samples in future investigations and forensic cases. This could prove vital within crime scene investigation and analysis as it could be improved further and used further within the forensic field.Key words: HPLC; Soil; Drugs; Trace Evidence; Forensic Science; Investigation.

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1. IntroductionDue to the heterogeneity, complexity, and transferability of soil, this makes it extremely valuable within forensic investigations as it can be analysed to compare unknown samples to known samples from crime scenes, and other locations that suspects could have transferred such evidence to or from. Forensic geoscience has become one of the most vital areas within forensic investigation and is a scientific discipline that applies the techniques developed to study earth materials pertaining to the law and has applications in any legal context (McCulloch et al., 2017). This means in cases where the materials of the earth can be used and may aid the investigators, along with the judges and the jurors of the court, to identify or gain an indication of what exactly happened, where and when the incident took place, and how or why the incident occurred. Due to the high transferability of soil, and its persistence and presence at a considerable amount of crime scenes, it can be used to aid within three major parts of the investigation, which are: corroborating with witness statements, the reconstruction of a crime, and the verification of suspect alibis.

Soil is classified as a geoforensic trace evidence material, and this means that it can be recovered from either the victim, the crime scene, or the suspect, along with any item that has transferred the material elsewhere, such as a car tires, bicycle tires, and shoes. This evidence can then be analysed and compared with control samples of soil to determine whether the material was transferred from locations that are of interest to forensic investigators.Since geoforensic science is a combination of forensic science and earth science, it is important to consider that the conceptual and pragmatic points of view both show distinct differences and it is vital to ensure meticulous considerations are given to significant differences so as to thoroughly and correctly decipher and explain the data that is generated using the chosen techniques. Such differences between the two include: the sample size and the level of spatial and temporal precision required, between the problems and questions encountered in forensic casework and those encountered in earth science research (McCulloch et al., 2017). Within forensic investigation, there are multiple techniques that were designed to analyse soil and other earth materials, all of which are aimed at the separation, identification and quantification of the materials at hand. For this study, HPLC was chosen as it was seen as the most effective and suitable technique available for analysing the soil samples retrieved from the four locations that were deemed to hold the most valuable evidence for the investigation that was taking place. This is because this technique could offer the opportunity to characterise and discriminate between the soil samples retrieved, whilst also being readily available to happen as the equipment needed was already available within the laboratories.

It has also been seen from other studies undertaken that this analytical technique has been deemed to show potential within the analysis of forensic soil samples.The aim of this experiment was to test a completely new analytical method to analyse and detect the presence of drugs within soil samples. The study also aimed to determine the effects of the soil itself upon the ability to detect the levels of the drug able to be analysed within the soil. Along with this, the study aimed to provide a new analytical method of detecting drugs within soil, that could be analysed with an easier implementable analysis method to retrieve data, whilst hoping to increase the conceivable occurrence of HPLC being used as a profiling tool with geoforensic samples in future criminal investigations.2.

Methodology2.1. Sample CollectionSamples of soil were collected from four separate locations within the borough of Liverpool. Location one (GP) was the garden of a lab assistant in the Wirral, location two (L3) was outside of the James Parsons building on Byrom Street, location three (WA3) was the garden of a lab assistant, and location four (Woodland) was a woodland area located behind the garden of a lab assistant. These sites were chosen due to their location within the borough, and these areas were more likely to have come into contact with the drugs in question due to the people within the area.The samples were taken from areas where the soil was exposed, as this soil was more forensically relevant due to the ease of transfer of the soil itself.

One sample was taken from each of the locations and were taken from one specific point at each. Once collected and separated from any gravel or grass present within the soil, each sample was placed and stored inside fully sealed plastic zip lock bags to protect them and prevent contamination occurring. 2.2. Sample Preparation – Spiked drug soil samplesThe samples were air-dried in a fume hood then kept in their plastic zip lock bags prior to the practical taking place.

Materials that were used during the experiment were; air-dried soil samples from specified locations, aspirin stock solution (1mg/ml), acetonitrile, a centrifuge and centrifuge tubes, a sonic bath, the Agilent 1100/1200 series HPLC UV instrument, HPLC tapered vials, a syringe and needle, and 0.45µm syringe filter. 250mg of the air-dried soil sample was weighed out four times and each of the four samples of soil were added to a 14ml centrifuge tube each, then 450µl of acetonitrile and 50µl of the aspirin (1mg/ml) stock solution was added to each of the centrifuge tubes.

The samples were mixed thoroughly using a vortex, before being sonicated for 15 minutes. Upon the completion of sonification, the solutions were then placed into the Hettich-Universal 320 centrifuge and were centrifuged for 20 minutes at 4000 RPM. Upon the completion of the centrifuge cycle, the samples were taken out and the supernatant was removed from each of the centrifuge tubes via a syringe and needle. The supernatant from each was then filtered via a 0.

45µm syringe filter into a HPLC tapered vial and proceeded to be analysed via the HPLC-UV instrument. 2.2.

1. Sample Preparation – Other samplesThe other samples were prepared in a very similar way, although the other samples did not need the aspirin stock solution adding to them, as they were not control samples, and were instead crime scene samples. 250mg of the air-dried soil from each location was weighed and added to 14ml centrifuge tubes, then 500µl of acetonitrile was added to each of the centrifuge tubes. Each sample was mixed thoroughly using a vortex, before also being sonicated for 15 minutes. Upon the completion of sonification, the samples were placed into the centrifuge for 20 minutes at 4000 RPM. Once the centrifuge cycle was complete, the samples were removed from the centrifuge and the supernatant was removed from each centrifuge tube with a syringe and needle. The supernatant from each sample was filtered into its own HPLC tapered vial via a 0.45µm syringe filter and proceeded to be analysed using the HPLC-UV instrument.

2.3. HPLC separation parametersThe samples were injected into the Agilent 1100 series HPLC-UV instrument, which had a column ODS of 15cm x 5micron, and a column temperature of 30oC.

The mobile phase was composed of UHQ water as mobile phase A, and a HPLC grade acetonitrile as mobile phase B. The flow rate of the HPLC-UV instrument was 1ml/min, with a run time of 35 minutes and a UV wavelength of 250nm. The table below shows the gradient run details of the HPLC-UV instrument, and the percentage of water to acetonitrile in comparison to each other (always adding up to 100% at each stage) as the time progressed.?Gradient time (min) % Water % ACN0.0 53 473.

0 45 5524.0 26 7429.0 2 9831.0 2 9832.0 53 4735.

0 53 47Table 1 – Gradient Run Details2.4. Data AnalysisThe data was analysed and HPLC chromatograms were produced from the Agilent OpenLab CDS Chemstation software, and this was also used to undertake integrations of the chromatograms. Each sample produced a variety of peaks, each of which had a diverse range of retention times in comparison to each other.

The data retrieved from each of the samples that were recovered from each of the locations would proceed to be analysed, and their peak retention times were compared to the peak retention times of the control sample to find similarities between them. If their peaks and retention times were similar between them, this would show that the crime scene samples did in fact contain aspirin, whereas if there were significant differences between them, then it would show that there was no aspirin present within the crime scene samples.3. Results and Discussion3.

1. HPLC ChromatogramsEach of the samples were ran through the Agilent OpenLab CDS Chemstation software and proceeded to produce chromatograms that were able undertake integrations also. These chromatograms were then able to be analysed in order to compare the peak retention times between the control sample, the drug spiked soil samples, and the blank soil samples to find out for definite whether the aspirin within the spiked soil samples was able to be detected using this method of analysis and detection. The chromatograms below (figures 1-9) show the results of the HPLC-UV analytical technique and how well it was able to detect the presence of the drugs within the spiked soul samples. They show the peak retention times of each sample, which were compared with each other to analyse whether the drugs in each sample were detected.

Figure 1 – Aspirin ChromatogramAs shown in the chromatogram above, there were only three peak retention times, the first of which was very tall compared to the other two peaks. With the use of this information, in comparison to the spiked soil samples and the blank soil samples, it became clear whether the aspirin in the soil samples was able to be detected using the HPLC-UV instrument, or whether it remained undetected and hidden by the soils own genealogy and biological factors. Figure 2 – Blank Soil Sample – GP Figure 3 – Spiked Soil Sample – GPAs could be seen in the above chromatograms, there were definite differences between the blank soil sample in comparison to the spiked soil sample, with more peaks being present within the second chromatograph. From these results, it was clear to see that there was a distinct presence and detection of the aspirin within the spiked soil sample from the GP location. This showed that the original genealogy of the soil itself had no effect on the aspirin stock solution, and the HPLC-UV instrument had no issue with detecting it within the samples. Figure 4 – Blank Soil Sample – L3 Figure 5 – Spiked Soil Sample – L3As with the first set of results, these chromatograms showed a distinct presence and detection of the aspirin within the soil. This could be seen with the larger number of peaks on the chromatogram produced from the spiked soil sample.

The peak retention time of the largest peak at the beginning of the chromatogram was in very close proximity of matching that of the aspirin chromatogram. It was clear to see the variety of differences between the peaks and their retention times on each of the chromatograms, as there were more on the spiked soil sample, but they were also more spread out than on the blank soil sample. Figure 6 – Blank Soil Sample – WA3 Figure 7 – Spiked Soil Sample – WA3Although the blank soil sample had slightly more peak retention times than the spiked soil sample, there was still a clear indication that the aspirin was present and detected within the spiked soil sample recovered from location WA3. As seen in previous results, the peak retention time of the very first peak on the spiked soil sample was in very close proximity of matching that of the aspirin stock solution, which provided evidence that the aspirin was clearly detected within the soil, and that the soil itself had no effect on the process. Figure 8 – Blank Soil Sample – Woodland Figure 9 – Spiked Soil Sample – WoodlandAs with the previous set of results, there were more peak retention times recorded on the blank soil sample than there were on the spiked soil sample, although this did not represent that the aspirin within the soil was not detected. In fact, regardless of this information, there was still a distinct and clear indication that the aspirin was present and detected within the soil sample.

This was indicated with the large peak at the earliest retention time as with the other results, as it was within very close proximity of matching that on the chromatogram of the aspirin stock solution.Each of the samples were easily distinguishable by their peak retention times in comparison to each other, and the spiked soil samples were easily distinguished by their large peaks at the earliest retention times on each chromatogram. Each chromatogram showed a distinct difference between the peaks of each sample, none of them being the same as each other, each of their retention times were completely different.

This demonstrated that even the soil samples themselves prior to being spiked were completely distinguishable in their retention times, most likely due to their original genealogy, but possibly due to the nature of the soil and how it had been treated prior to recovery at each location. With this information, it offered that the technique had the potential to be applied to further investigations within forensic science that involved that of earth materials and could possibly be used within many areas of the United Kingdom.Although, since there was only one sample of soil taken from each of the four locations, it was impossible to say whether the same results would have occurred with other samples from different sections of the location that the samples were recovered from, and there was no possible way of determining whether this technique could be a repeatable test, since there was only one sample of blank soil and one sample of spiked soil from each of the locations.

This demonstrated that a precautious approach to the interpretation of the results achieved from the samples when using HPLC-UV as an analytical technique for the detection of drugs in soil.?4. ConclusionIn conclusion, this study has displayed that the application of HPLC-UV as an analytical technique for the distinction of soil samples and the detection of drugs within soil samples was proven to be a valuable and advantageous technique, and favourable for future forensic use within crime scene investigation. Although there was only one blank sample and one spiked sample from each location, they still proved useful within the study as each of the spiked soil samples showed a presence of aspirin within it, in comparison to the blank soil samples. It is possible that with more samples from each location, and a repeatability test done on the extra samples, more reliable results would have been attained than what already was with the use of just one spiked sample from each location. Despite this being the case, the results that were attained were reliable and demonstrated what was hoped of the HPLC-UV analysis on each sample, and they showed that using this newly developed technique for this category of analysis established and identified a compelling opportunity for this strain of analysis to implemented for analysing geoforensic samples in further cases, and could benefit an amplitude of laboratories across the globe.The results of this study have thus indicated that it is feasible that HPLC analysis can be used as an effective technique when analysing and comparing different forensic soil samples retrieved and recovered from crime scene locations. It is also deemed from the results attained that HPLC analysis presents compelling potential to add to pre-existing techniques within geoforensic science, whilst also aiding with the investigation of crime scenes and the detection of the nature of the crime.

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